Graduation Term

2015

Degree Name

Master of Science (MS)

Department

School of Biological Sciences

Committee Chair

Viktor Kirik

Abstract

Plant cell walls are versatile structures, playing important roles in communication, defense, organization and support. The importance of each of these functions varies by cell type, with specialized cells often utilizing one or two functions more than others. Trichomes, or leaf hairs, and hypocotyl cells for instance, exhibit distinct cell wall characteristics. Trichomes have developed very thick cell walls with several raised structures, known as papillae, on their surfaces. It is believed that these cells function in defense against predators, making it difficult to crawl on the leaf surface, and in protection against ultra violet radiation, through refraction of light via the papillae. Hypocotyl cells on the other hand are long and narrow, with cell walls that undergo extensive remodeling to permit rapid oriented cell growth. This enables hypocotyl cells to elongate along the axis of plant development, pushing the developing shoot through the soil.

To better understand the molecular processes important for papillae deposition on the cell wall surface, as well as for cell wall building and cell elongation, we focused on the identification and characterization of the GLASSY HAIR (GLH) and SHORT HYPOCOTYL (SH) genes, in which mutants display underdeveloped papillae and small dark-grown hypocotyls, respectively.

A candidate gene, MED25, located within the mapping region on chromosome 1, was identified for glh1. The gene was sequenced, revealing a mutation site in the sixth intron. A genetic complementation test was conducted, which showed that MED25 was responsible for the glh1 glassy phenotype. A mutant rescue via transformation with the genomic region including MED25 was also performed, confirming mutation identity. The genetic introduction of markers for trichome development (GL2) and ethylene response (ETR2) into the mutant line demonstrated that the transcription of genes in trichomes is not generally changed in mutants. Î?-glucuronidase fusion with the promoter of the gene was also performed, confirming gene expression in trichomes.

The glh2 and glh3 mutations were rough mapped to chromosome 4 and 5, respectively. Genetic fine mapping was conducted to reduce the intervals in which the genes were thought to be located. Markers for trichome development were also introduced into each of these mutant lines to begin preliminary characterization of the nature of the mutations.

SH was rough mapped to a 2994kb region on chromosome 5. Known mutants within the mapping region that have sh-like phenotypes (cobra and procuste1/cesa6) were genetically or phenotypically tested to verify that sh was novel. A conditional radial expansion test revealed that sh does not exhibit a cobra phenotype. A confocal microscopy analysis of progeny from a sh x YFP-tagged CESA6 cross indicated that SH is not CESA6 and that CESA6 appears to be sequestered in Golgis of sh plants. In addition, we measured birefringence, an optical property of crystalline cellulose, in mutant and wild type trichomes, revealing a possible cellulose deficiency. We performed a biochemical cellulose assay to quantify the amount of cellulose in dark-grown mutant and wild type plants, confirming this possibility.

Access Type

Thesis-Open Access

DOI

http://doi.org/10.30707/ETD2015.Moore.C

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